A number of optical instruments require the splitting of light across a wide wavelength spectrum into a number of components corresponding to narrow wavelength bands. For example, fluorescent microscopes often incorporate light splitting technologies to distinguish between cells or cell types labeled with different fluorescent (FL) molecules. This is accomplished by subjecting the fluorescently labeled cells to one or more excitation wavelengths, such as from one or more lasers tuned to excite the particular fluorescent molecules, and filtering or separating the wavelength or wavelength range of interest from the entire emission spectrum for viewing. Traditionally, to achieve such wavelength separation, the light of emission spectrum (or simply, emission spectrum) is passed along multiple dichroic mirrors. As the light spectrum passes these serial dichroic mirrors, light within certain wavelength bands are separated out sequentially based on the properties of each dichroic mirror.
Similarly, in flow cytometry, biological cells or particles can be identified from one another in biological samples based on labeling with fluorescent molecules. The FL light is emitted from labeled cells as they pass through a laser beam in a flow channel travelling along a certain optical path. Since a cell may have a variety of markers indicative of its identity or status often two, three or more different fluorescent labels are used simultaneously requiring the instrument to have multi-color fluorescence detection capabilities. Separation and detection of these multi-color fluorescent wavelengths from the total emission spectra due to the multiple fluorescent labels is further complicated by additional measurements of forward scatter (FSC) and even more problematic, side scatter (SSC), which also require detection after exposure to the laser beam(s). Separation and detection of individual wavelengths or wavelength ranges within the emission spectrum is accomplished by directing the spectrum along a path where it is passed through a range of dichroic mirrors and filters, so that particular wavelength ranges are split sequentially from the wider emission spectrum and delivered to the appropriate light detectors. For example, cells of different lineages (e.g. blood cells) can often be distinguished from one another based on the presence of different protein markers (that may or may not be on the cell surface) and thus antibodies against these markers can be conjugated to different fluorescent molecules for their detection or measurement. When cells are labeled with antibodies conjugated to the popular fluorescent dyes such as fluorescein (FITC) and phycoerythrin (PE), each can be excited simultaneously, using a laser at 488 nm. In response, FITC emits FL light ranging from 470 to 610 nm, with a peak around 530 nm, whilst PE emits FL light ranging 530 to 660 nm, with a peak around 600 nm. Before separating and detecting the individual FITC and PE FL signals, the total FL light emitted from the cells is collected typically by an objective lens (single lens or a group of lenses) at 90° to the traveling direction of excitation laser. This total spectrum also includes the side scatter light at 488 nm. Once collected, separation and detection of side scatter as well as FITC and PE FL light can be accomplished as overviewed in FIG. 1 and as follows:                (a) The FL light coming from the objective lens is directed through, first, a 502 nm short-pass filter (i.e., passing light with wavelengths <502 nm whilst reflecting light >502 nm, wherein the passing light is then directed through a 488/10 nm band-pass filter to an optical detector such as photomultiplier tube (PMT) or photodiode for side scatter signal);        (b) The light reflected from the 502 nm short-pass filter is then directed through a second 556 nm long-pass filter (i.e. passing light with wavelengths >556 nm, whilst reflecting light <556 nm);        (c) The light reflected from the 556 nm long-pass filter is then directed through a 530/30 nm band-pass filter for an optical detector such as a PMT for FITC color detection;        (d) The light passing through the 556 nm long-pass filter is then directed through a 585/25 nm band-pass filter for an optical detector such as a PMT for PE color detection.        
The above approaches and other similar approaches require the use of multiple dichroic mirrors and band-filters for splitting light into its components having different, narrow wavelength bands. These approaches, whilst being adopted in many types of optical instruments, such as flow cytometers or some fluorescent microscopes, suffer from a number of limitations or shortcomings due to the use of a large number of optical components, such as: (1) high component cost; (2) lengthy and complicated optical alignment processes to achieve proper and efficient light splitting; and (3) large space requirements to cope with multiple components as well as their holders and to avoid spatial/mechanical interference between these components.
Thus, there remains a need to develop new optical light-splitting technologies which have simplified optical alignment procedures, occupy a small space and result in a lower cost.